The invention relates to treating various disorders associated with enhanced activity of kinase p38-xcex1. More specifically, it concerns piperadine and piperazine derivatives useful in these methods.
A large number of chronic and acute conditions have been recognized to be associated with perturbation of the inflammatory response. A large number of cytokines participate in this response, including IL-1, IL-6, IL-8 and TNF. It appears that the activity of these cytokines in the regulation of inflammation rely at least in part on the activation of an enzyme on the cell signaling pathway, a member of the MAP kinase family generally known as p38 and alternatively known as CSBP and RK. This kinase is activated by dual phosphorylation after stimulation by physiochemical stress, treatment with lipopolysaccharides or with proinflammatory cytokines such as IL-1 and TNF. Therefore, inhibitors of the kinase activity of p38 are useful anti-inflammatory agents.
Eye diseases associated with a fibroproliferative condition include retinal reattachment surgery accompanying proliferative vitreoretinopathy, cataract extraction with intraocular lens implantation, and post glaucoma drainage surgery.
PCT applications WO98/06715, WO98/07425, and WO 96/40143, all of which are incorporated herein by reference, describe the relationship of p38 kinase inhibitors with various disease states. As mentioned in these applications, inhibitors of p38 kinase are useful in treating a variety of diseases associated with chronic inflammation. These applications list rheumatoid arthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis and other arthritic conditions, sepsis, septic shock, endotoxic shock, Gram-negative sepsis, toxic shock syndrome, asthma, adult respiratory distress syndrome, stroke, reperfusion injury, CNS injuries such as neural trauma and ischemia, psoriasis, restenosis, cerebral malaria, chronic pulmonary inflammatory disease, silicosis, pulmonary sarcosis, bone resorption diseases such as osteoporosis, graft-versus-host reaction, Crohn""s Disease, ulcerative colitis including inflammatory bowel disease (IBD) and pyresis.
The above-referenced PCT applications disclose compounds which are p38 kinase inhibitors said to be useful in treating these disease states. These compounds are either imidazoles or are indoles substituted at the 3- or 4-position with a piperazine ring linked through a carboxamide linkage. Additional compounds which are conjugates of piperazines with indoles are described as insecticides in WO97/26252, also incorporated herein by reference.
Certain aroyl/phenyl-substituted piperazines and piperidines which inhibit p38-xcex1 kinase are described in PCT publication WO00/12074 published Mar. 9, 2000. In addition, indolyl substituted piperidines and piperazines which inhibit this enzyme are described in PCT publication No. WO99/61426 published Dec. 2, 1999. Carbolene derivatives of piperidine and piperazine as p38-xcex1 inhibitors are described in PCT/US00/07934 filed Mar. 24, 2000.
None of the foregoing patents describes the piperadine type derivatives described herein which specifically inhibit p38-xcex1.
The invention is directed to methods and compounds useful in treating conditions that are characterized by enhanced p38-xcex1 activity. These conditions include inflammation, proliferative diseases, and certain cardiovascular disorders as well as Alzheimer""s disease as further described below.
Compounds of the invention inhibit p38 kinase, the xcex1-isoform in particular, and are thus useful in treating diseases mediated by these activities. The compounds of the invention are of the formula (1): 
and the pharmaceutically acceptable salts thereof, or a pharmaceutical composition thereof, wherein:
Ar1 is an aryl group substituted with 0-5 non-interfering substituents, wherein two adjacent noninterfering substituents can form a fused aromatic or nonaromatic ring;
L1 and L2 are linkers;
each R1 is independently a noninterfering substituent;
Z1 is CR2 or N wherein R2 is hydrogen or a noninterfering substituent;
mis 0-4;
each of n and p is an integer from 0-2 wherein the sum of n and p is 0-3;
Ar2 is a substantially planar, monocyclic or polycyclic aromatic moiety having one or more optional ring heteroatoms, said moiety being optionally substituted with one or more non-interfering substituents, two or more of which may form a fused ring;
Z is xe2x80x94W1xe2x80x94COXjY wherein Y is COR3 or an isostere thereof; R3 is a noninterfering substituent, each of W and X is a spacer of 2-6 xc3x85, and each of i and j is independently 0 or 1;
wherein the smallest number of covalent bonds in the compound separating the atom of Ar1 bonded to L2 to the atom of Ar2 bonded to L1 is at least 6, where each of said bonds has a bond length of 1.2 to 2.0 angstroms; and/or wherein the distance in space between the atom of Ar1 bonded to L2 and the atom of Ar2 bonded to L1 is 4.5-24 angstroms;
with the proviso that the portion of the compound represented by Ar2xe2x80x94Z is not 
xe2x80x83wherein  represents a single or double bond; n is 0-3; one Z2 is CA or CRA and the other is CR, CR2, NR or N; A is xe2x80x94Wixe2x80x94COXjY wherein Y is COR or an isostere thereof, each of W and X is a spacer of 2-6 xc3x85, and each of i and j is independently 0 or 1; Z3 is NR or O; and each R is independently hydrogen or a noninterfering substituent.
The invention is further directed to methods of treating inflammation or proliferative conditions using these compounds. The invention is also directed to treating conditions associated with cardiac failure and Alzheimer""s disease using the invention compounds.
The compounds of formula (1) are useful in treating conditions which are characterized by overactivity of p38 kinase, in particular the xcex1-isoform. Conditions xe2x80x9ccharacterized by enhanced p38-xcex1 activityxe2x80x9d include those where this enzyme is present in increased amount or wherein the enzyme has been modified to increase its inherent activity, or both. Thus, xe2x80x9cenhanced activityxe2x80x9d refers to any condition wherein the effectiveness of these proteins is undesirably high, regardless of the cause.
The compounds of the invention are useful in conditions where p38-xcex1 kinase shows enhanced activity. These conditions are those in which fibrosis and organ sclerosis are caused by, or accompanied by, inflammation, oxidation injury, hypoxia, altered temperature or extracellular osmolarity, conditions causing cellular stress, apoptosis or necrosis. These conditions include ischemia-reperfusion injury, congestive heart failure, progressive pulmonary and bronchial fibrosis, hepatitis, arthritis, inflammatory bowel disease, glomerular sclerosis, interstitial renal fibrosis, chronic scarring diseases of the eyes, bladder and reproductive tract, bone marrow dysplasia, chronic infectious or autoimmune states, spinal chord injury and traumatic or surgical wounds. These conditions, of course, would be benefited by compounds which inhibit p38-xcex1. Methods of treatment with the compounds of the invention are further discussed below.
The compounds useful in the invention are derivatives of piperadine/piperazine-type compounds containing a mandatory substituent, Z attached to the aromatic moiety Ar2 The aromatic moiety is a substantially planar, monocyclic or polycyclic aromatic moiety having one or more optional ring heteroatoms. The aromatic moiety may be optionally substituted with one or more non-interfering substituents, two or more of which may form a fused ring.
In somewhat greater detail the aromatic moiety Ar2 comprises an optionally substituted monocyclic or polycyclic aromatic nucleus, wherein the aromatic nucleus consists of a carbocyclic or heterocyclic ring selected from (i) a five-membered heterocyclic or carbocyclic ring (ii) a six-membered carbocyclic or heterocyclic ring; (iii) a five-membered carbocyclic or heterocyclic ring fused to another five-membered carbocyclic or heterocyclic ring; (iv) a six-membered carbocyclic or heterocyclic ring fused to another six-membered carbocyclic or heterocyclic ring; and (v) a five-membered heterocyclic or carbocyclic ring fused to a six-membered carbocyclic or heterocyclic ring. Examples of the foregoing include the following aromatic moieties: 
where R is a noninterfering substituent.
Particular examples of Ar2 in formula (1) are such that the portion of compound (1) represented by L1xe2x80x94Ar2xe2x80x94Z is selected from the following: 
wherein n is 0, 1 or 2; X1 is NR, CR2, O or S; and each R is independently H or a noninterfering substituent; and two or more R groups may form a fused ring; 
wherein n is 0-4; R is H or a noninterfering substituent where two or more R groups may form a fused ring; and one or more ring carbons may be optionally replaced with nitrogen; 
wherein each n is inpendently 0 to 3; R is H or a noninterfering substituent, where two or more R groups may form a fused ring; and one or more ring carbons may be optionally replaced with nitrogen; 
wherein, subject to the proviso set forth above with respect to formula (1), one B is L1 and the other is Z; wherein a is 0 to 4 such that the positions on the six membered rings (1) and (3) to which (R)a is bonded can include X2 when X2 is C; b is 0-3 such that the positions on the five-membered rings (2) and (4) to which (R)b is bonded can include X2 and X1, when X2 is C and X1 is N or C; each X2 is independently N or CR; X1 is NR, CR2, O or S; each R is H or a noninterfering substituent where two or more R groups may form a fused ring; wherein one or more of the ring carbons that are at positions other than X2 or X1 and that are also not bound to B can be optionally replaced with N; 
wherein one B is L1 and the other is Z; a is 0-4 such that the positions on the rings (1) and (3) to which (R)a can be bonded include X2 and X1 where X2 is C and X1 is C or N; b is 0 or 3 such that the positions on the rings (2) and (4) to which (R)b can be bonded include X1, X2 and X3 when X1 is C or N and X2 and/or X3 are C; each X1 is independently NR, C(R)2, O or S; X2 and X3 are independently N or CR; each R is independently H or a noninterfering substituent where two or more R groups can optionally form a fused ring; wherein one or more of the ring carbons that are at positions other than X1, X2 or X3, and that are also not bound to B, can be optionally replaced with N.
Certain positions of the molecule of formula I are described as permitting xe2x80x9cnoninterfering substituents.xe2x80x9d This terminology is used because the substituents in these positions generally speaking are not relevant to the essential activity of the molecule taken as a whole. A wide variety of substituents can be employed in these positions, and it is well within ordinary skill to determine whether any particular arbitrary substituent is or is not xe2x80x9cnoninterfering.xe2x80x9d
As used herein, a xe2x80x9cnoninterfering substituentxe2x80x9d is a substituent which leaves the ability of the compound of formula (1) to inhibit p38-xcex1 activity qualitatively intact. Thus, the substituent may alter the degree of inhibition of p38-xcex1. However, as long as the compound of formula (1) retains the ability to inhibit p38-xcex1 activity, the substituent will be classified as xe2x80x9cnoninterfering.xe2x80x9d A number of assays for determining the ability of any compound to inhibit p38-xcex1 activity are available in the art. A whole blood assay for this evaluation is illustrated below. The gene for p38-xcex1 has been cloned and the protein can be prepared recombinantly and its activity assessed, including an assessment of the ability of an arbitrarily chosen compound to interfere with this activity. The essential features of the molecule are tightly defined. The positions which are occupied by xe2x80x9cnoninterfering substituentsxe2x80x9d can be substituted by conventional organic moieties as is understood in the art. It is irrelevant to the present invention to test the outer limits of such substitutions. The essential features of the compounds are those set forth with particularity herein.
In addition, L1 and L2 are described herein as linkers. The nature of such linkers is less important than the distance they impart between the portions of the molecule. Typical linkers include alkylene, i.e. (CH2)nxe2x80x94R; alkenylenexe2x80x94i.e., an alkylene moiety which contains a double bond, including a double bond at one terminus. Other suitable linkers include, for example, substituted alkylenes or alkenylenes, carbonyl moieties, and the like.
As used herein, xe2x80x9chydrocarbyl residuexe2x80x9d refers to a residue which contains only carbon and hydrogen. The residue may be aliphatic or aromatic, straight-chain, cyclic, branched, saturated or unsaturated. The hydrocarbyl residue, when so stated however, may contain heteroatoms over and above the carbon and hydrogen members of the substituent residue. Thus, when specifically noted as containing such heteroatoms, the hydrocarbyl residue may also contain carbonyl groups, amino groups, hydroxyl groups and the like, or contain heteroatoms within the xe2x80x9cbackbonexe2x80x9d of the hydrocarbyl residue.
As used herein, xe2x80x9cinorganic residuexe2x80x9d refers to a residue that does not contain carbon. Examples include, but are not limited to, halo, hydroxy, NO2, or NH2.
As used herein, the term xe2x80x9calkyl,xe2x80x9d xe2x80x9calkenylxe2x80x9d and xe2x80x9calkynylxe2x80x9d include straight- and branched-chain and cyclic monovalent substituents. Examples include methyl, ethyl, isobutyl, cyclohexyl, cyclopentylethyl, 2-propenyl, 3-butynyl, and the like. Typically, the alkyl, alkenyl and alkynyl substituents contain 1-10C (alkyl) or 2-10C (alkenyl or alkynyl). Preferably they contain 1-6C (alkyl) or 2-6C (alkenyl or alkynyl). Heteroalkyl, heteroalkenyl and heteroalkynyl are similarly defined but may contain 1-2 O, S or N heteroatoms or combinations thereof within the backbone residue.
As used herein, xe2x80x9cacylxe2x80x9d encompasses the definitions of alkyl, alkenyl, alkynyl and the related hetero-forms which are coupled to an additional residue through a carbonyl group.
The term xe2x80x9cAromaticxe2x80x9d with respect to moiety Ar1 refers to a monocyclic or fused bicyclic moiety such as phenyl or naphthyl; xe2x80x9cheteroaromaticxe2x80x9d also refers to monocyclic or fused bicyclic ring systems containing one or more heteroatoms selected from O, S and N. The inclusion of a heteroatom permits inclusion of 5-membered rings as well as 6-membered rings. Thus, typical aromatic systems include pyridyl, pyrimidyl, indolyl, benzimidazolyl, benzotriazolyl, isoquinolyl, quinolyl, benzothiazolyl, benzofuranyl, thienyl, furyl, pyrrolyl, thiazolyl, oxazolyl, imidazolyl and the like. Any monocyclic or fused ring bicyclic system which has the characteristics of aromaticity in terms of electron distribution throughout the ring system is included in this definition. Typically, the ring systems contain 5-12 ring member atoms.
Similarly, xe2x80x9carylalkylxe2x80x9d and xe2x80x9cheteroalkylxe2x80x9d refer to aromatic and heteroaromatic systems which are coupled to another residue through a carbon chain, including substituted or unsubstituted, saturated or unsaturated, carbon chains, typically of 1-6C. These carbon chains may also include a carbonyl group, thus making them able to provide substituents as an acyl moiety.
When the compounds of Formula (1) contain one or more chiral centers, the invention includes optically pure forms as well as mixtures of stereoisomers or enantiomers
With respect to the portion of the compound between the Ar1 and Ar2, linkers L2 and L1, in combination with the piperadine/piperazine ring, provide for separation of the atom of Ar1 bonded to L2 from the atom of Ar2 bonded to L1 by a defined minimum number of covalent bond lengths counted end-to-end through the compound, as opposed to a measurement of linear distance through space. More particularly, the smallest number of bonds counted end-to-end in the compound separating the atom of Ar1 bonded to L2 from the atom of Ar2 bonded to L1 is at least 5, and preferably from 6 to 12, wherein the length of each of such bonds is 1.2 to 2.0 angstroms. In terms of a linear distance through space, the linear distance measured through space from the atom of Ar1 bonded to L2 to the atom of Ar2 bonded to L1 is a distance of 4.5-24 xc3x85, preferably 6-20 xc3x85, and more preferably 7.5-10 xc3x85.
Typical, but nonlimiting, embodiments of L1 and L2 are CO and isosteres thereof, or optionally substituted isosteres, or longer chain forms. L2, in particular, may be alkylene or alkenylene optionally substituted with noninterfering substituents or L1 or L2 may be or may include a heteroatom such as N, S or O. Such substituents include, but are limited to, a moiety selected from the group consisting of alkyl, alkenyl, alkynyl, aryl, arylalkyl, acyl, aroyl, heteroaryl, heteroalkyl, heteroalkenyl, heteroalkynyl, heteroalkylaryl, NH-aroyl, halo, OR, NR2, SR, SOR, SO2R, OCOR, NRCOR, NRCONR2, NRCOOR, OCONR2, RCO, COOR, alkyl-OOR, SO3R, CONR2, SO2NR2, NRSO2NR2, CN, CF3, R3Si, and NO2, wherein each R is independently H, alkyl, alkenyl or aryl or heteroforms thereof, and wherein two substituents on L2 can be joined to form a non-aromatic saturated or unsaturated ring that includes 0-3 heteroatoms which are O, S and/or N and which contains 3 to 8 members or said two substituents can be joined to form a carbonyl moiety or an oxime, oximeether, oximeester or ketal of said carbonyl moiety.
Isosteres of CO and CH2, include SO, SO2, or CHOH, CO and CH2 are preferred. Thus, L2 is substituted with 0-2 substituents. Where appropriate, two optional substituents on L2 can be joined to form a non-aromatic saturated or unsaturated hydrocarbyl ring that includes 0-3 heteroatoms such as O, S and/or N and which contains 3 to 8 members. Two optional substituents on L2 can be joined to form a carbonyl moiety which can be subsequently converted to an oxime, an oximeether, an oximeester, or a ketal.
Ar1 is aryl, heteroaryl, including 6-5 fused heteroaryl, cycloaliphatic or cycloheteroaliphatic that can be optionally substituted. Ar is preferably optionally substituted phenyl.
Each substituent on Ar1 is independently a hydrocarbyl residue (1-20C) containing 0-5 heteroatoms selected from O, S and N, or is an inorganic residue. Preferred substituents include those selected from the group consisting of alkyl, alkenyl, alkynyl, aryl, arylalkyl, acyl, aroyl, heteroaryl, heteroalkyl, heteroalkenyl, heteroalkynyl, heteroalkylaryl, NH-aroyl, halo, OR, NR2, SR, SOR, SO2R, OCOR, NRCOR, NRCONR2, NRCOOR, OCONR2, RCO, COOR, alkyl-OOR, SO3R, CONR2, SO2NR2, NRSO2NR2, CN, CF3, R3Si, and NO2, wherein each R is independently H, alkyl, alkenyl or aryl or heteroforms thereof, and wherein two of said optional substituents on adjacent positions can be joined to form a fused, optionally substituted aromatic or nonaromatic, saturated or unsaturated ring which contains 3-8 members. More preferred substituents include halo, alkyl (1-4C) and more preferably, fluoro, chloro and methyl. These substituents may occupy all available positions of the aryl ring of Ar1, preferably 1-2 positions, most preferably one position. These substituents may be optionally substituted with substituents similar to those listed. Of course some substituents, such as halo, are not further substituted, as known to one skilled in the art.
Two substituents on Ar1 can be joined to form a fused, optionally substituted aromatic or nonaromatic, saturated or unsaturated ring which contains 3-8 members.
Between L1 and L2 is a piperidine-type moiety of the following formula: 
wherein Z1 is CR2 or N and R2 is H or a noninterfering substituent. Each of n and p is an integer from 0-2 wherein the sum of n and p is 0-3. The noninterfering substituents R2 include, without limitation, halo, alkyl, alkoxy, aryl, arylalkyl, aryloxy, heteroaryl, acyl, carboxy, or hydroxy. Preferably, R2 is H, alkyl, OR, NR2, SR or halo, where R is H or alkyl. Additionally, R2 can be joined with an R1 substituent to form an optionally substituted non-aromatic saturated or unsaturated hydrocarbyl ring which contains 3-8 members and 0-3 heteroatoms such as O, N and/or S. Preferred embodiments include compounds wherein Z1 is CH or N, and those wherein both n and p are 1.
R1 represents a noninterfering substituent such as a hydrocarbyl residue (1-20C) containing 0-5 heteroatoms selected from O, S and N. Preferably R1 is alkyl, alkoxy, aryl, arylalkyl, aryloxy, heteroalkyl, heteroaryl, heteroarylalkyl, RCO, xe2x95x90O, acyl, halo, CN, OR, NRCOR, NR, wherein R is H, alkyl (preferably 1-4C), aryl, or hetero forms thereof. Each appropriate substituent is itself unsubstituted or substituted with 1-3 substituents. The substituents are preferably independently selected from a group that includes alkyl, alkenyl, alkynyl, aryl, arylalkyl, acyl, aroyl, heteroaryl, heteroalkyl, heteroalkenyl, heteroalkynyl, heteroalkylaryl, NH-aroyl, halo, OR, NR2, SR, SOR, SO2R, OCOR, NRCOR, NRCONR2, NRCOOR, OCONR2, RCO, COOR, alkyl-OOR, SO3R, CONR2, SO2NR2, NRSO2NR2, CN, CF3, R3Si, and NO2, wherein each R is independently H, alkyl, alkenyl or aryl or heteroforms thereof and two of R1 on adjacent positions can be joined to form a fused, optionally substituted aromatic or nonaromatic, saturated or unsaturated ring which contains 3-8 members, or R1 is xe2x95x90O or an oxime, oximeether, oximeester or ketal thereof. R1 may occur m times on the ring; m is an integer of 0-4. Preferred embodiments of R1 comprise alkyl (1-4C) especially two alkyl substituents and carbonyl. Most preferably R1 comprises two methyl groups at positions 2 and 5 or 3 and 6 of a piperidinyl or piperazinyl ring or xe2x95x90O preferably at the 5-position of the ring. The substituted forms may be chiral and an isolated enantiomer may be preferred.
Z is xe2x80x94Wixe2x80x94COXjY wherein Y is COR3 or an isostere thereof and R3 is a noninterfering substituent. Each of W and X is a spacer and may be, for example, optionally substituted alkyl, alkenyl, or alkynyl, each of i and j is 0 or 1. Preferably, W and X are unsubstituted. Preferably, j is 0 so that the two carbonyl groups are adjacent to each other. Preferably, also, i is 0 so that the proximal CO is adjacent the ring. However, compounds wherein the proximal CO is spaced from the ring can readily be prepared by selective reduction of an initially glyoxal substituted Ar2.
The noninterfering substituent represented by R3, when R3 is other than H, is a hydrocarbyl residue (1-20C) containing 0-5 heteroatoms selected from O, S and/or N or is an inorganic residue. Preferred are embodiments wherein R3 is H, or is straight or branched chain alkyl, alkenyl, alkynyl, aryl, arylalkyl, heteroalkyl, heteroaryl, or heteroarylalkyl, each optionally substituted with halo, alkyl, heteroalkyl, SR, OR, NR2, OCOR, NRCOR, NRCONR2, NRSO2R, NRSO2NR2, OCONR2, CN, COOR, CONR2, COR, or R3Si wherein each R is independently H, alkyl, alkenyl or aryl or the heteroatom-containing forms thereof, or wherein R3 is OR, NR2, SR, NRCONR2, OCONR2, or NRSO2NR2, wherein each R is independently H, alkyl, alkenyl or aryl or the heteroatom-containing forms thereof, and wherein two R attached to the same atom may form a 3-8 member ring and wherein said ring may further be substituted by alkyl, alkenyl, alkynyl, aryl, arylalkyl, heteroalkyl, heteroaryl, heteroarylalkyl, each optionally substituted with halo, SR, OR, NR2, OCOR, NRCOR, NRCONR2, NRSO2R, NRSO2NR2, OCONR2, or R3Si wherein each R is independently H, alkyl, alkenyl or aryl or the heteroatom-containing forms thereof wherein two R attached to the same atom may form a 3-8 member ring, optionally substituted as above defined.
Other preferred embodiments of R3 are H, heteroarylalkyl, xe2x80x94NR2, heteroaryl, xe2x80x94COOR, xe2x80x94NHRNR2, heteroaryl-COOR, heteroaryloxy, xe2x80x94OR, heteroaryl-NR2, xe2x80x94NROR and alkyl. Most preferably R3 is isopropyl piperazinyl, methyl piperazinyl, dimethylamine, piperazinyl, isobutyl carboxylate, oxycarbonylethyl, morpholinyl, aminoethyldimethylamnine, isobutyl carboxylate piperazinyl, oxypiperazinyl, ethylcarboxylate piperazinyl, methoxy, ethoxy, hydroxy, methyl, amine, aminoethyl pyrrolidinyl, aminopropanediol, piperidinyl, pyrrolidinyl-piperidinyl, or methyl piperidinyl.
Isosteres of COR3 as represented by Y are defined as follows. The isosteres have varying lipophilicity and may contribute to enhanced metabolic stability. Thus, Y, as shown, may be replaced by the isosteres in Table 1.
Thus, isosteres include tetrazole, 1,2,3-triazole, 1,2,4-triazole and imidazole.
The compounds of formula (1) may be supplied in the form of their pharmaceutically acceptable acid-addition salts including salts of inorganic acids such as hydrochloric, sulfuric, hydrobromic, or phosphoric acid or salts of organic acids such as acetic, tartaric, succinic, benzoic, salicylic, and the like. If a carboxyl moiety is present on the compound of formula (1), the compound may also be supplied as a salt with a pharmaceutically acceptable cation.
Synthesis of the Invention Compounds
Copending, commonly-assigned U.S. Ser. No. 09/575,060, incorporated herein by reference in its entirety, illustrated the following reaction scheme for conversion of a 4-benzyl piperidinyl-indole-5-carboxamide to the glyoxalic acid compounds of the invention and derivatives thereof: 
In the present invention, the indole moiety is generalized to Ar2 in formula (1) above where Ar2 is a substantially planar, monocyclic or polycyclic aromatic moiety having one or more optional ring heteroatoms, said moiety being optionally substituted with one or more non-interfering substituents, two or more of which may form a fused ring. Preferably the moiety Ar2 comprises an optionally substituted monocyclic or polycyclic aromatic nucleus, wherein said aromatic nucleus consists of carbocyclic or heterocyclic ring selected from (i) a five-membered heterocyclic or carbocyclic ring (ii) a six-membered carbocyclic or heterocyclic ring; (iii) a five-membered carbocyclic or heterocycloc ring fused to another five-membered carbocyclic or heterocyclic ring; (iv) a six-membered carbocyclic or heterocyclic ring fused to another six-membered carbocyclic or heterocyclic ring; and (v) a five-membered heterocyclic or carbocyclic ring fused to a six-membered carbocyclic or heterocyclic ring. Formula (1), as required by the proviso stated above, excludes the indole type compounds disclosed and claimed in U.S. Ser. No. 09/575,060 filed May 21, 1999 and incorporated herein by reference.
As disclosed commonly assigned in U.S. Ser. No. 09/575,060, the glyoxal type substituent at position 3 can be generalized to xe2x80x94WiCOXjY.
The Ar2 moiety may be generalized as: 
Methods to synthesize the compounds of the invention are, in general, known in the art. For example, commonly assigned U.S. Ser. No. 09/575,060, incorporated herein by reference in its entirety, disclosed that piperidine moieties can be obtained using the following reaction scheme 
where an appropriate piperidone such as I, is treated with substituted benzyl phosphonate esters in the presence of a base such as sodium hydride to give alkenes which can be reduced to the corresponding substituted 4-benzylpiperidine such as II. The hydrogenations are typically done in the presence of catalytic metals in solvents such as methanol, ethanol and ethyl acetate.
An alternative to the above disclosed in U.S. Ser. No. 09/575,060 as follows: 
where isonipecotoyl chlorides such as I can be used to acylate appropriately substituted benzenes (ArH) in the presence of a Lewis acid such as aluminum chloride to give the ketones II. Further modifications of the carbonyl moiety of II using methods and routes generally known can then lead to the desired compounds III.
The following reaction schemes illustrate methods for preparing compounds of the present invention. 
Assays for p38 xcex1 Kinase Inhibition
For each of the assay procedures described below, the TNF-xcex1 production correlates to the activity of p38-xcex1 kinase.
A. Human Whole Blood Assay for p38 Kinase Inhibition
Venous blood is collected from healthy male volunteers into a heparinized syringe and is used within 2 hours of collection. Test compounds are dissolved in 100% DMSO and 1 xcexcl aliquots of drug concentrations ranging from 0 to 1 mM are dispensed into quadruplicate wells of a 24-well microtiter plate (Nunclon Delta SI, Applied Scientific, So. San Francisco, Calif.). Whole blood is added at a volume of 1 ml/well and the mixture is incubated for 15 minutes with constant shaking (Titer Plate Shaker, Lab-Line Instruments, Inc., Melrose Park, Ill.) at a humidified atmosphere of 5% CO2 at 37xc2x0 C. Whole blood is cultured either undiluted or at a final dilution of 1:10 with RPMI 1640 (Gibco 31800+NaHCO3, Life Technologies, Rockville, Md. and Scios, Inc., Sunnyvale, Calif.). At the end of the incubation period, 10 xcexcl of LPS (E. coli 0111:B4, Sigma Chemical Co., St. Louis, Mo.) is added to each well to a final concentration of 1 or 0.1 xcexcg/ml for undiluted or 1:10 diluted whole blood, respectively. The incubation is continued for an additional 2 hours. The reaction is stopped by placing the microtiter plates in an ice bath and plasma or cell-free supernates are collected by centrifugation at 3000 rpm for 10 minutes at 4xc2x0 C. The plasma samples are stored at xe2x88x9280xc2x0 C. until assayed for TNF-xcex1 levels by ELISA, following the directions supplied by Quantikine Human TNF-xcex1 assay kit (RandD Systems, Minneapolis, Minn.).
IC50 values are calculated using the concentration of inhibitor that causes a 50% decrease as compared to a control.
B. Enriched Mononuclear Cell Assay for p38 Kinase Inhibition
The enriched mononuclear cell assay, the protocol of which is set forth below, begins with cryopreserved Human Peripheral Blood Mononuclear Cells (HPBMCs) (Clonetics Corp.) that are rinsed and resuspended in a warm mixture of cell growth media. The resuspended cells are then counted and seeded at 1xc3x97106 cells/well in a 24-well microtitre plate. The plates are then placed in an incubator for an hour to allow the cells to settle in each well.
After the cells have settled, the media is aspirated and new media containing 100 ng/ml of the cytokine stimulatory factor Lipopolysaccharide (LPS) and a test chemical compound is added to each well of the microtiter plate. Thus, each well contains HPBMCs, LPS and a test chemical compound. The cells are then incubated for 2 hours, and the amount of the cytokine Tumor Necrosis Factor Alpha (TNF-xcex1) is measured using an Enzyme Linked Inmunoassay (ELISA). One such ELISA for detecting the levels of TNF-xcex1 is commercially available from RandD Systems. The amount of TNF-xcex1 production by the HPBMCs in each well is then compared to a control well to determine whether the chemical compound acts as an inhibitor of cytokine production.
LPS Induced Cytokine Synthesis in HPBMCs
Cryopreserved HPBMC (cat #CC-2702 Clonetics Corp)
LGM-3 media (cat #CC-3212 Clonetics Corp)
LPS stock 10 xcexcg/ml (Cat. No. L 2630 serotype 0111:B4 Sigma)
Human TNF-xcex1 ELISA (RandD Systems)
DNase I (10 mg/ml stock)
Preparation of Cells
LGM-3 media warmed to 37xc2x0 C.
5 xcexcl of DNase I stock added to 10 ml media.
Cells thawed rapidly and dispersed into above.
Centrifuge 200xc3x97gxc3x9710 min @ RT.
Pellet up in 10 ml sterile PBS.
Centrifuge 200xc3x97gxc3x9710 min @ RT.
Pellet resuspended in 10 ml LGM-3 then diluted to 50 ml with LGM-3.
Perform cell count.
Adjust to 1xc3x97E06 cells/well.
Seed 1 ml/well of a 24 well plate.
Place plate in incubator to plate down for 1 hour.
Preparation of Incubation Media
LGM-3 containing 100 ng/ml LPS (e.g. 50 ml media plus 0.5 ml LPS stock) Aliquot into 2 ml aliquots and add 1000xc3x97inhibitor dilutions.
Incubation
When cells have plated down aspirate media away and overlay with 1 ml relevant incubation media. Return plate to incubator for 2 hours or 24 hours. Remove supernatants after incubation to a labeled tube and either perform TNF (or other) ELISA immediately or freeze for later assay.
IC50 values are calculated using the concentration of inhibitor that causes a 50% decrease as compared to a control.
Administration and Use
The compounds of the invention are useful among other indications in treating conditions associated with inflammation. Thus, the compounds of formula (1) or their pharmaceutically acceptable salts are used in the manufacture of a medicament for prophylactic or therapeutic treatment of mammals, including humans, in respect of conditions characterized by excessive production of cytokines and/or inappropriate or unregulated cytokine activity on such cells as cardiomyocytes, cardiofibroblasts and macrophages.
The compounds of the invention inhibit the production of cytokines such as TNF, IL-1, IL-6 and IL-8, cytokines that are important proinflammatory constituents in many different disease states and syndromes. Thus, inhibition of these cytokines has benefit in controlling and mitigating many diseases. The compounds of the invention are shown herein to inhibit a member of the MAP kinase family variously called p38 MAPK (or p38), CSBP, or SAPK-2. The activation of this protein has been shown to accompany exacerbation of the diseases in response to stress caused, for example, by treatment with lipopolysaccharides or cytokines such as TNF and IL-1. Inhibition of p38 activity, therefore, is predictive of the ability of a medicament to provide a beneficial effect in treating diseases such as Alzheimer""s, coronary artery disease, congestive heart failure, cardiomyopathy, myocarditis, vasculitis, restenosis, such as occurs following coronary angioplasty, atherosclerosis, IBD, rheumatoid arthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis and other arthritic conditions, multiple sclerosis, acute respiratory distress syndrome (ARDS), asthma, chronic obstructive pulmonary disease (COPD), silicosis, pulmonary sarcosis, sepsis, septic shock, endotoxic shock, Gram-negative sepsis, toxic shock syndrome, heart and brain failure (stroke) that are characterized by ischemia and reperfusion injury, surgical procedures, such as transplantation procedures and graft rejections, cardiopulmonary bypass, coronary artery bypass graft, CNS injuries, including open and closed head trauma, inflammatory eye conditions such as conjunctivitis and uveitis, acute renal failure, glomerulonephritis, inflammatory bowel diseases, such as Crohn""s disease or ulcerative colitis, graft vs. host disease, bone resorption diseases like osteoporosis, type II diabetes, pyresis, psoriasis, cachexia, viral diseases such as those caused by HIV, CMV, and Herpes, and cerebral malaria.
Within the last several years, p38 has been shown to comprise a group of MAP kinases designated p38-xcex1, p38-xcex2, p38-xcex3 and p38-xcex4. Jiang, Y., et al., J. Biol Chem (1996) 271:17920-17926 reported characterization of p38-xcex2 as a 372-amino acid protein closely related to p38-xcex1. In comparing the activity of p38-xcex1 with that of p38-xcex2, the authors state that while both are activated by proinflammatory cytokines and environmental stress, p38-xcex2 was preferentially activated by MAP kinase kinase-6 (MKK6) and preferentially activated transcription factor 2, thus suggesting that separate mechanisms for action may be associated with these forms.
Kumar, S., et al., Biochem Biophys Res Comm (1997) 235:533-538 and Stein, B., et al., J Biol Chem (1997) 272:19509-19517 reported a second isoform of p38-xcex2, p38-xcex22, containing 364 amino acids with 73% identity to p38-xcex1. All of these reports show evidence that p38-xcex2 is activated by proinflammatory cytokines and environmental stress, although the second reported p38-xcex2 isoform, p38-xcex22, appears to be preferentially expressed in the CNS, heart and skeletal muscle compared to the more ubiquitous tissue expression of p38-xcex1. Furthermore, activated transcription factor-2 (ATF-2) was observed to be a better substrate for p38-xcex22 than for p38-xcex1, thus suggesting that separate mechanisms of action may be associated with these forms. The physiological role of p38-xcex21 has been called into question by the latter two reports since it cannot be found in human tissue and does not exhibit appreciable kinase activity with the substrates of p38-xcex1.
The identification of p38-xcex3 was reported by Li, Z., et al., Biochem Biophys Res Comm (1996) 228:334-340 and of p38-xcex4 by Wang, X., et al., J Biol Chem (1997) 272:23668-23674 and by Kumar, S., et al., Biochem Biophys Res Comm (1997) 235:533-538. The data suggest that these two p38 isoforms (xcex3 and xcex4) represent a unique subset of the MAPK family based on their tissue expression patterns, substrate utilization, response to direct and indirect stimuli, and susceptibility to kinase inhibitors.
Various results with regard to response to drugs targeting the p38 family as between p38-xcex1 and either the putative p38-xcex21 or p38-xcex22 or both were reported by Jiang, Kumar, and Stein cited above as well as by Eyers, P. A., et al., Chem and Biol (1995) 5:321-328. An additional paper by Wang, Y., et al., J Biol Chem (1998) 273:2161-2168 suggests the significance of such differential effects. As pointed out by Wang, a number of stimuli, such as myocardial infarction, hypertension, valvular diseases, viral myocarditis, and dilated cardiomyopathy lead to an increase in cardiac workload and elevated mechanical stress on cardiomyocytes. These are said to lead to an adaptive hypertrophic response which, if not controlled, has decidedly negative consequences. Wang cites previous studies which have shown that in ischemia reperfusion treated hearts, p38 MAPK activities are elevated in association with hypertrophy and programmed cell death. Wang shows in the cited paper that activation of p38-xcex2 activity results in hypertrophy, whereas activation of p38-xcex1 activity leads to myocyte apoptosis. Thus, selective inhibition of p38-xcex1 activity as compared to p38-xcex2 activity will be of benefit in treating conditions associated with cardiac failure. These conditions include congestive heart failure, cardiomyopathy, myocarditis, vasculitis, vascular restenosis, valvular disease, conditions associated with cardiopulmonary bypass, coronary artery bypass, grafts and vascular grafts. Further, to the extent that the xcex1-isoform is toxic in other muscle cell types, xcex1-selective inhibitors would be useful for conditions associated with cachexia attributed to TNF or other conditions such as cancer, infection, or autoimmune disease.
Thus, the invention encompasses the use of compounds which selectively inhibit the activity of the p38-xcex1 isoform for treating conditions associated with activation of p38-xcex1, in particular those associated with cardiac hypertrophy, ischemia or other environmental stress such as oxidation injury, hyperosmolarity or other agents or factors that activate p38-xcex1 kinase, or cardiac failure, for example, congestive heart failure, cardiomyopathy and myocarditis.
The manner of administration and formulation of the compounds useful in the invention and their related compounds will depend on the nature of the condition, the severity of the condition, the particular subject to be treated, and the judgement of the practitioner; formulation will depend on mode of administration. As the compounds of the invention are small molecules, they are conveniently administered by oral administration by compounding them with suitable pharmaceutical excipients so as to provide tablets, capsules, syrups, and the like. Suitable formulations for oral administration may also include minor components such as buffers, flavoring agents and the like. Typically, the amount of active ingredient in the formulations will be in the range of 5%-95% of the total formulation, but wide variation is permitted depending on the carrier. Suitable carriers include sucrose, pectin, magnesium stearate, lactose, peanut oil, olive oil, water, and the like.
The compounds useful in the invention may also be administered through suppositories or other transmucosal vehicles. Typically, such formulations will include excipients that facilitate the passage of the compound through the mucosa such as pharmaceutically acceptable detergents.
The compounds may also be administered topically, for topical conditions such as psoriasis, or in formulation intended to penetrate the skin. These include lotions, creams, ointments and the like which can be formulated by known methods. The compounds may also be administered by injection, including intravenous, intramuscular, subcutaneous or intraperitoneal injection. Typical formulations for such use are liquid formulations in isotonic vehicles such as Hank""s solution or Ringer""s solution.
Alternative formulations include nasal sprays, liposomal formulations, slow-release formulations, and the like, as are known in the art.
Any suitable formulation may be used. A compendium of art-known formulations is found in Remington""s Pharmaceutical Sciences, latest edition, Mack Publishing Company, Easton, Pa. Reference to this manual is routine in the art.
The dosages of the compounds of the invention will depend on a number of factors which will vary from patient to patient. However, it is believed that generally, the daily oral dosage will utilize 0.001-100 mg/kg total body weight, preferably from 0.01-50 mg/kg and more preferably about 0.01 mg/kg-10 mg/kg. The dose regimen will vary, however, depending on the conditions being treated and the judgment of the practitioner.
It should be noted that the compounds of formula (1) can be administered as individual active ingredients, or as mixtures of several embodiments of this formula. In addition, the inhibitors of p38 kinase can be used as single therapeutic agents or in combination with other therapeutic agents. Drugs that could be usefully combined with these compounds include natural or synthetic corticosteroids, particularly prednisone and its derivatives, monoclonal antibodies targeting cells of the immune system, antibodies or soluble receptors or receptor fusion proteins targeting immune or non-immune cytokines, and small molecule inhibitors of cell division, protein synthesis, or MRNA transcription or translation, or inhibitors of immune cell differentiation or activation.
As implied above, although the compounds of the invention may be used in humans, they are also available for veterinary use in treating animal subjects.
The following examples are intended to illustrate but not to limit the invention. The compounds described and prepared in examples 1-4 below are inhibitors of p38-xcex1 kinase.